Macromolecular MRI contrast agents based upon dendrimers obviate many of the deficiencies of serum albumin or linear polymer based MRI contrast agents of comparable size. This is due to the iterative synthesis by which they are created that promotes controlled size and shape of the dendrimer concomitantly generating the means for reproducible chemistry key to the clinical translation of such agents. To create MRI contrast agents with dendrimers, the terminal primary amines of dendrimers are modified with chelated Gd(III) technology developed in our laboratories. Historically, these reagents demonstrated a molar relaxivity 6 times that of Magnevist, the currently approved MRI contrast agent. Excellent conventional whole body MR imaging and 3D T-O-F MR angiograms have been obtained using PAMAM and polypropyleneimine dendrimer-based agents.Past studies established that macromolecular chelate conjugated dendrimer based Gd(III) MR contrast agents can be tuned for various applications by adjusting fundamental criteria: generation (MW & size), core elements (lipophilicity & charge), PEG conjugation (prolong circulation and minimize liver & other organ uptake), lysine co-administration (renal clearance), and conjugation to targeting vectors (molecular targeting). These dendrimer based agents have also been selectively targeted, not only by conjugation to antibodies, but by other vectors, such as avidin to deliver exceptionally high levels of Gd(III) into disseminated intraperitoneal ovarian cancer tumor. This was done in conjunction with an optical imaging agent in parallel with our creation of multi-modality dendrimer based imaging agents. The incorporation of a NIR optical imaging dye into the MRI agent added an enhanced level of sensitivity to complement the resolution of the MRI imaging and provided an additional level of sensitivity for the imaging of lymphatics and sentinel nodes that can be envisioned as being translated to an intraoperative scenario wherein MRI imaging and mapping would supplement real-time surgical intervention and excision of malignancy. While the chemistry established the ability to create such macromolecular agents, the imaging resulted in compromised targeting which defined that these agents require very careful systematic investigation combined with equally careful defined characterization. New chelation chemistry for conjugation of Gd(III) complexes to dendrimer has been prompted by the need to re-invent this field moving it from aqueous chemistry to organic phase solvents to enhance both characterization and consistency of yields. This chemistry has also evolved specialized analogs of established bifunctional chelation agents (patent filed) to address development of site-specific conjugation chemistry required for actively targeted dendrimer based imaging agents e.g, maleimides targeting a unique thiol residue, or agents functionalized with alkyne or azide groups for click chemistry conjugation strategies.The impact of NFS related Gd(III) toxicity resultant from use of less than adequately stable MRI contrast agents combined with low renal excretion had prompted a complete halt of projects with application of new directionality in the choice of bifunctional chelating agent. Redirection towards the use of DOTA as the bifunctional chelating agent, however, with the Gd(III) complex pre-formed prior to conjugation to all targeting vectors has been demonstrated to successfully traverse toxicity concerns. This effort was put into place with all of the MR contrast projects fully migrated to the exclusive use of a pre-complexation of the Gd(III) conjugate strategy using DOTA to also eliminate characterization complexity resulting from the creation of exceedingly difficult to characterize mixtures of products that limited reproducibility which would have complicated clinical translation of these agents.Results from the studies validate that this transformation not only can be a successful strategy despite warnings of decreased solubility (not true), but that far greater molar relaxivity can be achieved by this means. We have reported a 5-fold enhancement over the prior technology while concurrently decreasing the actual physical amount of Gd(III) conjugated to the dendrimer by 65% further increasing the safety margin. The impact of this result reaches across to all macromolecular MR contrast agents regardless of platform to fully address safety, characterization, and reproducibility thereby furthering an entire fields potential for clinical translation of such agents. The exquisite advantages of the dendrimer based agents over low molecular weight agents continues to be very clearly demonstrated.In parallel to this improvement to abrogate toxicity concerns that has resulted in superior dendrimer based agents, the ability to finally move forward with actively targeted dendrimer based contrast agents that are discrete characterized agents was achieved through conjugation of a cystamine core dendrimer derived dendron conjugated in a 1:1 form with an antibody fragment. While successful, both isolation and purification were challenging with the dendrimer generation employed as well as relatively lower relaxivity results indicating the need to move onwards to a higher dendrimer generation. This investigation continues in parallel with the creation of a PEG appended trifunctional imaging agent that will permit incorporation of a radiological probe (PET imaging) with an optical probe.Patent applications continue to be prosecuted in parallel to publications revealing this technology to the field which should prove to make it yet more valuable to HHS and should also contribute to translation of this technology into the clinic. Studies of MRI and other imaging modality agents in collaboration with the Molecular Imaging Program have unfortunately been terminated due to a lack of cooperation and access to instrumentation residing therein in what was to be a resource for all NCI researchers despite agreements to the contrary. However, established collaborations with Radiology, CC, the PET Dept, CC, NIMH, and extramural researchers at Johns Hopkins continue to be fruitful.